Objective: To study the permeability of leflunomide and the effect of liposome on its permeability. Methods: Modified Franz diffusion cell adopted as apparatus for in vitro skin permeation. Various excised skins were used as permeation barriers and permeation coefficient was used as index. The concentration of leflunomide in the samples was measured by HPLC. Results: Leflunomide had a good permeability, but the effect of liposome on permeability was not very strong. Conclusion: It is feasible that leflunomide is applied through skin. [

Objective To study the relationship between Oxfordshire Community Stroke Project(OCSP)classification and middle cerebral artery(MCA)lesions detected by digital subtraction angiography among patients with acute MCA territorial infarction.Methods One hundred and fifty-four patients with acute MCA territorial infarction,who were obtained from Jinling Hospital during May 2002 to December 2005,were classified into total anterior circulation infarction(TACI),partial anterior circulation infarction(PACI)and lacunar infarction(LACI).Results Compared with LACI subtype,prevalence of MCA main stem occlusion in patients with TACI subtype was higher(P

OBJECTIVE:To identify the influence of twisting or untwisting Taixi(K13) on the brain function by observing functional mag-netic resonance imaging.METHODS:Twelve healthy young cases were enrolled in this research.A block design including three blocks which baseline and stimulation appear alternately was adapted.In stimulation phase,the needle was twisted manually,or not twisted.Scanned images were analyzed using SPM2.RESULTS:The right superior temporal gyrus(BA22),left medial frontal gyrus(BA46),followed by right and left postcentral gyrus of parietal lobe(BA2,BA3),left inferior frontal gyrus(BA45) and left inferior parietal lobule(BA40) were evoked by acu-puncture at Taixi,but there was no activeation area after untwisting acupoint.CONCLUSION:The different areas evoked by twisting or untwisting Taixi acupoint are closely related with the internal organs as well as the course of the channels and collaterals.

OBJECTIVE:To establish the RP-HPLC method for determination of methyl hydroxybenzoate and ethyl hydroxybenzoate in oral liquid preparations METHODS:The Nova-Park C18 column(3 9mm?150mm,5?m)was used The mobile phase consisted of acetonitrile-water(25∶75)at a flow rate of 1ml/min,the detection wavelength was set at 254nm and the sensitivity was set at 0 01 AUFS RESULTS:The linear ranges of methyl hydroxybenzoate and ethyl hydroxybenzoate were 0 4 412～13 2 360?g/ml(r=0 9 999) and 0 6 672～20 0 160?g/ml(r=0 9 998)respectively CONCLUSION:The method is simple,rapid and accurate It can be used to detect antiseptics in hospital preparations

Objective To study the effects of CD2-associated protein (CD2AP) on podocyte adhesion and extension ability and to explore its possible mechanism. Methods Conditionally immortalized murine podocyte cell line was cultured in RPMI 1640 medium at 33℃permissive conditions. The podocytes were transfected with CD2AP small interfering RNA (siRNA) and serambing sequences labeled with fluorescein were taken as control. The transfected podocytes were trypsinized and seed into collagen IV coated plates. The relative cell adhesion and cell area were examined 90 min later. Apoptotic rates of CD2AP siRNA transfected podoeytes and different PAN concentrations incubated podoeytes were detected by flow cytometer. The distribution of F-actin was observed under laser scanning confoeal microscope. Nephrin protein expression and its phosphorylation level were examined by immunofluorescence and Western blot. Results The relative ceil adhesion of CD2AP siRNA transfected podocytes was apparently lower than that of control group[(41.72±6.07)% vs (64.46±8.53)%, P<0.05]. The cell area analysis had the similar result. The apoptotic rate of CD2AP siRNA transfected podocytes was significantly higher than that of the controls [(5.73±0.61)% vs (3.26±0.45)%, P<0.05]. 100 mg/L PAN could markedly induce podocytes to apoptosis and impair cell adhesion ability (P<0.05). Nevertheless, no significant difference was found in cell body spreading (P>0.05). The distribution of F-actin in CD2AP depletion podocytes was apparently altered. The expression of nephrin protein and its phosphorylation level was conspicuously descended to some degree (P<0.05). Conclusions CD2AP depletion facilitates podocyte apoptosis and impairs cell adhesion function. Cytoskeleton confusion and nephrin signaling weakness caused by CD2AP depletion may he partly responsible for the decline of cell adhesion and spreading.

AIM: To investigate the effect of glibenclamide (Glib) on the viability and acid-base equilibrium of glioblastoma cells.METHODS: U251 cells and U87 cells were treated with Glib at different concentrations.The inhibitory rates were detected by CCK-8 assay.The effective dose was screened and the experiment was divided into control group and drug treatment groups.The migration ability was monitored by wound healing assay, and intracellular pH was detected by pH indicator fluorescent probe.The protein expression levels of inwardly-rectifying potassium channel 4.1 (Kir4.1) and monocarboxylate transport protein 1 (MCT1) were determined by Western blot.RESULTS: The half maximal inhibitory concentrations (IC50) of Glib for 48 h exposure of U251 cells and U87 cells were 400.20 μmol/L and 553.70 μmol/L, respectively.The effective inhibition doses of Glib for U251 cells were from the ranges of 100 μmol/L to 1 600 μmol/L, and those for U87 cells were from 50 μmol/L to 1 600 μmol/L in a concentration-dependent manner (P<0.05).Glib not only inhibited the migration (P<0.05) of U251 cells and U87 cells, which was negatively correlated with drug concentration (P<0.05), but also reduced the intracellular fluorescence intensity in experimental group (P<0.05), suggesting that with the increase in drug concentration, the intracellular pH decreased gradually (P<0.05).The protein expression of Kir4.1 and MCT1 was down-regulated by treatment with Glib, and was negatively correlated with concentration of Glib.CONCLUSION: Glib, a kind of potassium channel blocker, induces intracellular acidification via down-regulating the expression of Kir4.1 and MCT1, thus inhibiting the growth of glioblastoma in a certain dose range.

Objective To verify the relationship between transforming growth factor-β1(TGF-β1) and interleukin-10 (IL-10) in breast milk and allergic diseases development in infants. Methods Totally 191 mothers (99 allergics and 92 controls) and their full-term newborns participated in this prospective study on development of children atopy. Maternal blood, cord blood, colostrum and mature milk were assayed for TGF-β1 and IL-10 by enzyme-linked immunosorbent assay. Infants underwent pediatrician evaluation for allergic diseases at six months old. Concentrations of TGF-β1 and IL-10 from allergic and non-allergic mothers and prevalence of allergic diseases of infants were compared. Results The level of IgE in allergic mothers was 30 750 IU/L(6600-410000 IU/L),lower than that in non-allergic mothers[50000 IU/L(7100-610000 IU/L)](Z=-3. 444,P=0. 001).No difference in the concentration of TGF-β1, IL-10 and IgE in mature milk was observed between allergic and non-allergic mothers. TGF-β1, IL-10 and IgE levels in colostrum of allergic mothers were 2300 pg/ml(620-7000 pg/ml), 12. 8 pg/ml(7.5-560.0 pg/ml)and 7000 IU/L(5100-56000 IU/L),significantly higher than those in non-allergic mothers[1830 pg/ml(1240-9400 pg/ml), 11. 1 pg/ml (7. 2-630.0 pg/ml)and 6700 IU/L(5200-35000 IU/L)] (Z=-2. 215, -2. 730 and -2. 706,P＜0.05).In both allergic and non-allergic mothers, TGF-β1 and IL-10 levels in cord blood were higher than those in maternal blood, while IgE was lower. TGF-βl and IL-10 and IgE levels in colostrum were higher than mature milk(P＜0.05). At six months old, the prevalence of allergic diseases of infants from allergic mothers(59. 6%, 59/99) was significantly higher than those from non-allergic mothers (21. 7%, 20/92)(x2= 28. 177, P= 0. 000). The prevalence of allergic diseases of infants who completed two weeks' colostrum-fed after birth (44.5 %, 73/164) was significantly higher than those who did not (22.2%,6/27)(x2 =4. 749,P-=0. 029). Conclusions High concentration of TGF-βl and IL-10 in colostrum does not show any protective effect against allergic diseases in infants. The prevalence of allergic diseases of colostrum-fed infants is significantly higher than non colostrum-fed infants, showing that colostrum-fed might play a role in allergic diseases development.

AIM: To observe the effect of Tripterygium glycosides on NOXs-ROS-NLRP3 inflammatory signa-ling pathways in the colon tissue in dextran sulphate sodium (DSS)-induced ulcerative colitis (UC) mice, and to investi-gate the underlying mechanisms.METHODS: BALB/c mice were used and the mouse model of UC was established by DSS induction.The mice were randomly divided into 5 groups (model group, low-, medium-and high-dose Tripterygium glycosides groups, and normal group).The colon tissues were collected 21 d after Tripterygium glycosides gavage.The mRNA expression of NLRP3, ASC and caspase-1 in the colon tissues was detected by real-time PCR.The caspase-1 ex-pression in the colorectal mucosa was observed by immunohistochemical method.ELISA was used to detect the protein le-
vels of IL-1α, TNF-αand IL-13.The production of reactive oxygen species (ROS) was measured by chemiluminescence technique, and the consumption rate of NADPH, which was inhibited by DPI, was analyzed to determine the activity of NADPH oxidases (NOXs).The neutrophils were isolated, and the ROS production, NOXs activity, and the mRNA ex-pression of NLRP3, ASC and caspase-1 were also detected.RESULTS: The colon tissues were abnormal with different de-grees in Tripterygium glycosides groups, and histopathological scores were lower than that in model group.In Tripterygium glycosides groups, in addition to the mRNA expression levels of caspase-1 in the colon tissues between normal group and high-dose group, ROS production, NOXs activity and the mRNA expression levels of NLRP3, ASC and caspase-1 in the colon tissues and colon-isolated neutrophils were lower than those in model group (P <0.05), and higher than those in normal group (P <0.05).The results of pairwise comparison for the efficacy of Tripterygium glycosides administration showed that the above indexes were statistically significant except the mRNA expression levels of caspase-1 between middle-dose group and high-dose group.Tripterygium glycosides administration significantly decreased the expression levels of proinflammatory cytokines IL-1αand TNF-αin the homogenates of colon tissues in the model mice (P <0.05).No differ-ence of IL-13 expression among the groups was observed.CONCLUSION: Tripterygium glycosides inhibits NOXs-ROS-NLRP3 inflammatory signaling pathways to reduce the expression of IL-1α, TNF-αand other proinflammatory cytokines, and attenuates DSS-induced ulcerative colitis in mice, by which the neutrophils might be involved in the process.

Objective: To explore mechanism of fasudil combined salmeterol and fluticasone propionate powder for inhalation (Seretide) in treating chronic obstructive pulmonary disease (COPD) complicated pulmonary artery hypertension (PAH).Methods: A total of 120 patients accorded with diagnostic standards of COPD and PAH, who hospitalized in our department from Jan 2013 to Oct 2014, were selected.According to random number table method, patients were randomly and equally divided into routine treatment group, fasudil group (received intravenous drip of fasudil based on routine treatment group) and combined treatment group (received additional Seretide therapy based on fasudil group).Levels of nitric oxide (NO), endothelin-1 (ET-1), C reactive protein (CRP) and erythrocyte sedimentation rate (ESR) were measured and compared among three groups before and after treatment.Results: Compared with before treatment, after two-week treatment, there were significant reductions in levels of CRP, ESR and ET-1, and significant rise in NO level in three groups, P<0.05 or<0.01;compared with routine treatment group after treatment, there were significant reductions in levels of CRP[(14.8±3.3) mg/L vs.(12.9±3.6) mg/L vs.(11.4±3.4) mg/L], ESR[(37.3±8.9) mm/h vs.(32.9±8.8) mm/h vs.(29.3±5.6) mm/h]and ET-1[(63.1±11.2) ng/L vs.(57.5±8.1) ng/L vs.(53.1±8.9) ng/L], and significant rise in NO level[(70.2±10.7) μmol/L vs.(76.0±8.0) μmol/L vs.(80.5±11.3) μmol/L]in fasudil group and combined treatment group, P<0.05 or<0.01;compared with fasudil group, there were significant reductions in levels of CRP, ESR and ET-1, and significant rise in NO level in combined treatment, P<0.05 all.Conclusion: Fasudil hydrochloride combined Seretide can significantly reduce levels of ESR, CRP and ET-1, and increase NO level in COPD + PAH patients.It may improve prognosis in these patients, which is worth extending.

Objective To study the effect of overexpression of TRPC6 on Ang Ⅱ-induced apoptosis of mouse podocytes in vitro and to explore the possible mechanisms. Methods Mouse TRPC6 cDNA eukaryotie expression vector pEGFP-NI-mTRPC6 was transfected to conditionally immortalized routine podocyte cell line by liposome. The fluorescent microscopy was used to examine the expression of EGFP after 24 hours. The change of TRPC6 protein expression was observed by Western-blot. Podocytes were treated by different concentrations of Ang Ⅱ. The podocyte intracellular calcium concentration was measured with laser-scanning con_focal microscope. The expression of Bax and Bcl-2 mRNA was assessed by RT-PCR and the expression of Bax and Bcl-2 protein was measured by Western-blot. The apoptotic ratio of podocytes was monitored by flow cytometry and Hoechst staining. Results About 35% of the cells expressed EGFP. An up-regulation of protein expression of TRPC6 was detected in podocytes when transfected with pEGFP-N1-mTRPC6 (P<0.01). The overexpression of TRPC6 promoted the Ang Ⅱ-induced influx of extracellular calcium and elevated the expression of Bax but decreased the expression of Bcl-2 (P<0.01, P<0.05). The apoptotic ratio of podocyte was (2.50±0.72)% when treated by low-dose Ang Ⅱ (10-10 mol/L), and it was increased to (4.33±0.45)% when transfected with pEGFP-N1-mTRPC6 (P <0.05 ). Transfection with pEGFP-NI-mTRPC6 increased apoptosis rate from (15.46± 1.40)% to (18.33±0.87)%(P<0.01) by high-dose Ang Ⅱ (10-6 mol/L). Conclusion TRPC6 plays an important role in the Ang Ⅱ-induced apoptosis of podocytes by promoting the influx of extraeellular calcium, which leads to the apoptosis cascade initiation.